大肠埃希菌感染对U937细胞凋亡的影响及机制探讨

目的探讨大肠埃希菌(E.coli)感染对人巨噬细胞系U937细胞的促凋亡作用及其机制。方法调整U937细胞与E.coli浓度比为120时,放入培养基中培养0、10、20、30、60和90 min,获得感染的U937细胞。采用流式细胞仪检测不同感染时间U937细胞的细胞凋亡率,Western blot法检测U937细胞胞浆内第2线粒体激活剂(Smac)和X连锁凋亡抑制蛋白(XIAP)的表达水平。在E.coli感染前60 min用浓度0、10、20和40μmol.L-1的Em-belin预处理U937细胞,然后在E.coli诱导U937细胞凋亡30 min后收集细胞,用Annexin V FITC/PI双染后用流式细胞仪分析U937细胞的凋亡情况。结果当U937细胞E.coli浓度比为120时,感染0、10、20、30、60和90 min时U937细胞凋亡率分别为(3.02±0.78)%、(6.67±1.34)%、(10.56±1.02)%、(33.92±2.66)%、(46.98±3.12)%和(69.02±4.69)%,呈时间依赖性。U937细胞感染E.coli后发生凋亡,并且凋亡率随着E.coli感染时间的延长而升高。Smac的表达随着E.coli感染时间的延长逐渐升高,XIAP的表达则随着感染时间的延长而逐渐降低,而且不同感染时间组比较差别有统计学意义(P<0.05)。加入XIAP的抑制剂Embelin后,U937细胞的凋亡率随着Embelin的浓度增加而逐渐升高(P<0.05)。结论人巨噬细胞系U937细胞感染E.coli后发生凋亡,其凋亡的发生与影响Smac和XIAP的表达有关。Embelin通过特异性地抑制XIAP表达,增加E.coli诱导的U937细胞凋亡率。

Mechanism and effect of Escherichia coli infection on apoptosis of U937 cell

Objective To explore the mechanism and effect of Escherichia coli（E.coli） infection on apoptosis of U937 cell in human macrophagic system.Methods U937 cells and E.coli were putted into culture media 0,10,20,30,60 and 90 minutes when the concentration ratio of them was adjusted to 120,then the infected U937 cells were found.The rates of U937 apoptosis in different time were detected by the flow cytometer.The expressive levels of caspase second mitochondrial activator factor（Smac） in cytoplasm of U937 cells and X chromosome linked inhibitor of apoptosis protein（XIAP） were detected by Western blot.U937 cells were pretreated by Embelin which concentration were 0,10,20 and 40 μmol·L-1 at sixty minutes before the E.coli infection,and then U937 cells were gathered at thirty minutes after apoptosis induced by E.coli.The U937 cells apoptosis were analyzed by flow cytometer after Annexin V FITC/PI double-dyeing.Results When U937 cellsE.coli equaled to 120,the apoptosis rates of U937 cells showed time-dependent,the apoptosis rates at 0,10,20,30,60 and 90 minutes after infection were（3.02±0.78）%,（6.67±1.34）%,（10.56±1.02）%,（33.92±2.66）%,（46.98±3.12）% and（69.02±4.69）%.U937 cells began to apoptosis after infected by E.coli,and the apoptosis rate was increased as the infected time extended.With the extension of E.coli infection′s time,the expression of Smac was raised gradually,whereas,the expression of XIAP was decreased gradually.Moreover,there were significant differences among different infected time（P0.05）.The apoptosis rate of U937 cells was increased gradually with the increasing of Embelin′s concentration after putting into inhibitor Embelin（P0.05）.Conclusion U937 cells in human macrophagic system began to apoptosis after infected by E.coli,and the occurrence and the influence of apoptosis were concerned with expression of Smac and XIAP.Through inhibit the expression of XIAP specifically,the Embelin increased the apoptosis rate of U937 cells after induced by E.coli.